Establishment of reference intervals for plasma protein electrophoresis and comparison of biochemical and protein electrophoresis evaluation of albumin in central bearded dragons (Pogona vitticeps).

The aim of this study is to develop a reference interval for plasma protein electrophoresis for wild free-roaming central bearded dragons (Pogona vitticeps). This will assist the veterinarian in diagnosing and monitoring diseases in wild and captive animals. The study group consisted of 29 wild central bearded dragons of which 22 were males and 7 were females. The plasma samples were obtained from animals in a previous study and were stored frozen at -20°C for 2-3 years before analysis. Plasma protein electrophoresis was performed on 100 uL of plasma. The data were evaluated for the difference between reproductively active females and pooled males with non-reproductively active females as well as the difference between biochemical and protein electrophoresis albumin. Biochemical albumin was found to be significantly higher than electrophoresis albumin in all animals, with a mild proportional bias present. Plasma protein electrophoresis identified albumin, α - 1 and 2, ß - 1 and 2 and γ fractions. The reproductively active females in this study exhibited results that were distinct from pooled males and non-reproductively active females. This study provides a reference interval for plasma protein electrophoresis in central bearded dragons that can be used as a tool for health and disease monitoring and diagnosis. The importance of the use of method and sex/season appropriate reference intervals in reptile medicine is confirmed in this study.

Haematology and plasma biochemistry reference intervals in wild bearded dragons (Pogona vitticeps).

The aim of the study is to provide haematology and plasma biochemistry reference intervals for wild free-roaming central bearded dragons (Pogona vitticeps). These data will aid the veterinarian in determining the health and husbandry status of the pet bearded dragon. The study group consisted of 130 wild central bearded dragons. The data were assigned into two groups: Breeding Season (BS) from September to November and Non-Breeding Season (NBS) from December to March. These groups were further divided into sex and reproduction status with females determined to either be gravid or non-gravid. The 83 male and 47 female dragons were captured in north western New South Wales, Australia, centred around the towns of Bourke, Wanaaring and Tibooburra. Haematological and plasma biochemistry parameters were measured using standard methods. Data were analysed for the effects of sex, season and gravid status. There were significant differences between sexes in PCV, basophils, albumin, globulin, amylase, calcium, phosphate and triglycerides. There were seasonal differences in PCV, basophils, uric acid, glucose, globulin, calcium, phosphate, cholesterol and triglycerides. There were significant differences in phosphate and cholesterol between gravid and non-gravid dragons within the breeding season. There were variations in haematology and biochemistry parameters depending on sex, season and reproductive status of females. These reference intervals differed from previous studies of captive central bearded dragons and can provide some insight into captive conditions compared to wild animals.

Oral progestin induces rapid, reversible suppression of ovarian activity in the cat.

The influence of oral progestin (altrenogest; ALT) on cat ovarian activity was studied using non-invasive fecal steroid monitoring. Queens were assigned to various ALT dosages: (1) 0mg/kg (control; n=5 cats); (2) 0.044 mg/kg (LOW; n=5); (3) 0.088 mg/kg (MID; n=6); or (4) 0.352 mg/kg (HIGH; n=6). Fecal estrogen and progestagen concentrations were quantified using enzyme immunoassays for 60 days before, 38 days during and 60 days after ALT treatment. Initiation of follicular activity was suppressed in all cats during progestin treatment, whereas controls continued to cycle normally. Females (n=6) with elevated fecal estrogens at treatment onset completed a normal follicular phase before returning to baseline and remained suppressed until treatment withdrawal. All cats receiving oral progestin re-initiated follicular activity after treatment, although MID cats experienced the most synchronized return (within 10-16 days). Mean baseline fecal estrogens and progestagens were higher (P<0.05) after treatment in HIGH, but not in LOW or MID cats compared to pre-treatment values. The results demonstrate that: (1) oral progestin rapidly suppresses initiation of follicular activity in the cat, but does not influence a follicular phase that exists before treatment initiation; and (2) queens return to normal follicular activity after progestin withdrawal. This study provides foundational information for research aimed at using progestin priming to improve ovarian response in felids scheduled for ovulation induction and assisted breeding.

Advances in reproductive science for wild carnivore conservation.

Knowledge about reproduction is critical for predicting the viability of wildlife populations in nature and for managing breeding programmes in captivity. Intensive species-based studies are the priority, because reproductive mechanisms are extraordinarily diverse, even within the same taxonomic family. Carnivores deserve more attention as such species are highly vulnerable to environmental change and human persecution. The present review provides contemporary illustrations of how reproductive science is contributing to understand unique reproductive mechanisms that are both of fundamental and applied interest. In the case of the endangered African wild dog (Lycaon pictus) free-living in South Africa, non-invasive faecal corticosteroid assessments have yielded new insights about the impact of animal relocation and reintroduction on adaptive responses, reproductive fitness and survival. For the maned wolf (Chrysocyon brachyurus), advances have been made in characterizing and comparing reproductive traits in free-ranging vs captive individuals. For the cheetah (Acinonyx jubatus), recent studies have focused on the cryosensitivity of sperm and the ability to develop a field-friendly sperm cryo-method. The by-product has been a large-scale frozen repository of sperm from wild-caught cheetahs useful for infusing new genes into ex situ populations. Finally, rigorous, multi-disciplinary and cross-institutional reproductive studies of the black-footed ferret (Mustela nigripes), including the use of artificial insemination, have contributed to the remarkable recovery and restoration of this species, once on the brink of extinction. In summary, advances in reproductive science are not necessarily related to 'assisted breeding'. However, understanding the unique ways of carnivore reproduction greatly contributes to species management and conservation.

Slow cooling prevents cold-induced damage to sperm motility and acrosomal integrity in the black-footed ferret (Mustela nigripes).

The endangered black-footed ferret (Mustela nigripes) has benefited from artificial insemination; however, improved sperm cryopreservation protocols are still needed. The present study focused on identifying factors influencing gamete survival during processing before cryopreservation, including: (1) the presence or absence of seminal plasma; (2) temperature (25 degrees C v. 37 degrees C); (3) type of medium (Ham's F10 medium v. TEST yolk buffer [TYB]); (4) cooling rate (slow, rapid and ultra-rapid); and (5) the presence or absence of glycerol. Seminal plasma did not compromise (P > 0.05) sperm motility or acrosomal integrity. Sperm motility traits were maintained longer (P < 0.05) at 25 degrees C than at 37 degrees C in Ham's or TYB, but temperature did not affect (P > 0.05) acrosomal integrity. Overall, TYB maintained optimal (P < 0.05) sperm motility compared with Ham's medium, but Ham's medium maintained more (P < 0.05) intact acrosomes than TYB. Slow cooling (0.2 degrees C min(-1)) was optimal (P < 0.05) compared to rapid cooling (1 degrees C min(-1)), and ultra-rapid cooling (9 degrees C min(-1)) was found to be highly detrimental (P < 0.05). Results obtained in TYB with 0% or 4% glycerol were comparable (P > 0.05), indicating that 4% glycerol was non-toxic to ferret sperm; however, glycerol failed to ameliorate the detrimental effects of either rapid or ultra-rapid cooling. The results of the present study demonstrate that the damage observed to black-footed ferret spermatozoa is derived largely from the rate of cooling.

A Retinal Prosthesis Technology Based on CMOS Microelectronics and Microwire Glass Electrodes.

A very large format neural stimulator device, to be used in future retinal prosthesis experiments, has been designed, fabricated, and tested. The device was designed to be positioned against a human retina for short periods in an operating room environment. Demonstrating a very large format, parallel interface between a 2-D microelectronic stimulator array and neural tissue would be an important step in proving the feasibility of high resolution retinal prosthesis for the blind. The architecture of the test device combines several novel components, including microwire glass, a microelectronic multiplexer, and a microcable connector. The array format is 80 times 40 array pixels with approximately 20 microwire electrodes per pixel. The custom assembly techniques involve indium bump bonding, ribbon bonding, and encapsulation. The design, fabrication, and testing of the device has resolved several important issues regarding the feasibility of high-resolution retinal prosthesis, namely, that the combination of conventional CMOS electronics and microwire glass provides a viable approach for a high resolution retinal prosthesis device. Temperature change from power dissipation within the device and maximum electrical output current levels suggest that the device is acceptable for acute human tests.

Short term suppression of follicular recruitment and spontaneous ovulation in the cat using levonorgestrel versus a GnRH antagonist.

Suppression and subsequent rebound of ovarian activity using a progestin (levonorgestrel; Norplant) versus a GnRH antagonist (antide) was assessed in the domestic cat via fecal estradiol and progesterone metabolite analyses. Following an initial dose-response trial, queens were assigned to one of four treatments: (1) antide, two 6 mg/kg injections 15 days apart (n = 8 cats); (2) levonorgestrel, six silastic rods (36 mg levonorgestrel/rod) implanted for 30 days (n = 8); (3) control injections (n = 5); and (4) control implants (n = 5). Steroid metabolites were quantified from daily fecal samples for 90 days before, 30 days during, and 90 days after treatment. Antide and levonorgestrel inhibited estrous cyclicity in contrast to continued cyclicity in controls. Cats already at estradiol baseline in antide (n = 7) and levonorgestrel (n = 4) groups remained inhibited during treatment. In females with elevated estradiol levels at treatment onset (Day 0), a normal estradiol surge was completed before concentrations declined to baseline (approximately Days 5-7) and remained suppressed throughout the remaining treatment period. Additionally, 56% of treatment animals exhibited at least one spontaneous ovulation during the pre-treatment period, but no female ovulated during treatment with levonorgestrel or antide. Antide-treated cats exhibited lower (P < 0.05) baseline estradiol concentrations during treatment compared to pre- and post-treatment. In contrast, levonorgestrel induced elevations in baseline estradiol following treatment compared to pre- and during treatment intervals. Control females showed no change (P > 0.05) in baseline estradiol throughout the study period. All levonorgestrel and antide cats returned to estrus after treatment withdrawal. Results demonstrate that: (1) both antide and levonorgestrel are effective for inducing short-term suppression of follicular recruitment and ovulation in the cat; (2) inhibition is reversible; and (3) GnRH antagonists and progestins differentially regulate basal estradiol secretion. This study also confirmed a relatively high incidence of spontaneous ovulation in the cat, a species generally considered to be an induced ovulator.

Acrosomal integrity and capacitation are not influenced by sperm cryopreservation in the giant panda.

Sperm cryopreservation and artificial insemination are important management tools for giant panda breeding and the preservation of extant genetic diversity. This study examined the influence of freeze-thawing on sperm function, specifically capacitation. Sperm from nine giant pandas were assessed before and after rapid (- 40 and - 100 degrees C/min) cryopreservation by incubation in HEPES-buffered Ham's F10 medium with and without the capacitation accelerators, 3-isobutyl-1-methylxanthine (IBMX) and dibutyryl cyclic AMP (dbcAMP). At 0, 3 and 6 h of exposure, aliquots were assessed for sperm motility traits and capacitation, defined as the proportion of sperm with intact acrosomes following exposure to solubilised zonae pellucidae (ursid or felid) or calcium ionophore subtracted from the proportion of sperm with intact acrosomes before exposure. Although mean+/-S.E.M. sperm motility post-thaw (56.1 +/- 3.9% at 0 h) was less (P < 0.05) than pre-freeze (71.7 +/- 6.0%), there was no difference (P > 0.05) in the proportion of acrosome-intact sperm (fresh, 93.0 +/- 1.7% versus cryopreserved-thawed, 81.7 +/- 4.7% at 0 h). Incidence of capacitation was greater (P < 0.05) in fresh sperm incubated with capacitation accelerators IBMX and dbcAMP (9 h: 50.9 +/- 1.1) compared with fresh sperm incubated without accelerators (9 h: 41.2 +/- 1.1%). Frozen-thawed sperm preincubated without accelerators underwent capacitation (49.6 +/- 1.1%) to a greater extent (P < 0.05) compared with these fresh counterparts. Thawed samples with (9 h: 45.9 +/- 1.4%) and without accelerators (9 h: 41.2 +/- 1.1%) did not differ (P > 0.05) during the 9-h incubation. We conclude that giant panda spermatozoa (1) undergo capacitation in vitro with or without chemical accelerators and (2) withstand a rapid cryopreservation protocol, including retaining normal acrosomal integrity and functional capacitation ability.

Linkage of reproductive sciences: from 'quick fix' to 'integrated' conservation.

Our laboratory has experienced four phases in understanding how the reproductive sciences contribute to genuine conservation of biodiversity. The first is the 'quick fix phase' in which the erroneous assumption is made that extant knowledge and techniques are readily adaptable to an unstudied wild animal to produce offspring rapidly. The second is the 'species-specificity phase' in which it is recognized that every species has evolved unique reproductive mechanisms that must be mastered before propagation can be enhanced. The third is the 'applicability phase' in which one grasps that all the new knowledge and technology are of minimal relevance without the cooperation of wildlife managers. The final phase is 'integration', the realization that reproduction is only one component in an abundantly complex conservation puzzle that requires interweaving many scientific disciplines with elaborate biopolitical, economic and habitat variables. These phases are illustrated using 20 years of experience with wildlife species, including the cheetah, black-footed ferret and giant panda. We conclude that the foremost value of the reproductive sciences for conserving endangered species is the discipline's powerful laboratory tools for understanding species-specific reproductive mechanisms. Such scholarly information, when applied holistically, can be used to improve management by natural or, occasionally, assisted breeding. Genuine conservation is achieved only when the reproductive knowledge and technologies are integrated into multidisciplinary programmes that preserve species integrity ex situ and preferably in situ.

The phenomenon and significance of teratospermia in felids.

The common domestic cat is an important research model for endangered felids, as well as for studying genetic dysfunctions, infectious diseases and infertility in humans. Especially significant is the trait of teratospermia (ejaculation of < 40% morphologically normal spermatozoa) that commonly occurs in about 70% of the felid species or subspecies studied to date. Teratospermia, discovered more than two decades ago in the cheetah, is important: (i) for understanding the significance of sperm form and function; and (ii) because this condition is common in human males. It is apparent from IVF that deformed spermatozoa from teratospermic felids do not fertilize oocytes. However, the inability of spermatozoa from teratospermic males to bind, penetrate and decondense in the cytoplasm of the oocyte is not limited to malformed cells alone. Normal shaped spermatozoa from teratospermic males have reduced functional capacity. IVF results have consistently revealed a direct correlation between teratospermia and compromised sperm function across felid species and populations. The most significant differences between normospermic (> 60% normal spermatozoa per ejaculate) and teratospermic felids include: (i) the time required for sperm capacitation and the acrosome reaction to occur in vitro; (ii) culture media requirements for capacitation in vitro; (iii) phosphorylation patterns of tyrosine residues on sperm membrane proteins during capacitation; (iv) susceptibility to chilling-induced sperm membrane damage; (v) sensitivity to osmotic stress; (vi) stability of sperm DNA; (vii) sperm protamine composition; and (viii) fertilizing ability after intracytoplasmic sperm injection. In conclusion, (i) the felids (including wild species) are valuable for studying the functional significance of both pleiomorphic and normally formed spermatozoa from teratospermic donors, and (ii) the impact of teratospermia is expressed at both macrocellular and subcellular levels.

Understanding the basic reproductive biology of wild felids by monitoring of faecal steroids.

The ability to track gonadal activity is essential for understanding the fundamentals of reproduction. Faecal steroid metabolite monitoring is a well established tool for evaluating reproductive processes in diverse mammalian species, including felids. Domestic cats were used as a model and injection of radiolabelled oestradiol, progesterone, testosterone and cortisol revealed that > 85% of metabolites were excreted in faeces with a time lag of 12-24 h. Steroids were extracted by boiling faecal material (wet or dry) in 90% aqueous ethanol followed by immunoassay with group-specific antibodies that crossreact with excreted metabolites. This approach was used to illustrate the diversity of oestrous cycle characteristics, gonadal responses to photoperiod and ovulatory sensitivity within the felid taxon. Longitudinal analyses demonstrated that faecal oestradiol increases with observed oestrus, and that the duration of the oestrous cycle varies among felid species. Seasonality in gonadal activity was observed in some species (for example clouded leopard, Pallas' cat), whereas other species (for example margay, cheetah, oncilla) are cyclic all year round. Although cats are considered induced ovulators, non-mating and spontaneous ovulation occurred in some species (for example domestic cat, clouded leopard, lion, leopard, margay) with varying frequency. There was also evidence that suppressed ovarian activity and oestrus occurred in group-housed cats (for example cheetahs). As assisted reproductive techniques, such as artificial insemination, are becoming increasingly important for managing zoo species, steroid metabolite monitoring has been especially useful for examining the efficacy of associated hormonal therapies. Exogenous gonadotrophins used to induce ovulation often caused ovarian hyperstimulation, which resulted in a maternal endocrine environment that differed from that of naturally mated cats. Finally, there is evidence that the adrenal status of animals managed under different husbandry conditions can be assessed non-invasively, thereby enhancing our understanding of how social and environmental factors affect animal well-being and reproductive fitness. In summary, understanding the basic endocrinology of endangered felids generates knowledge that can be used to improve management strategies. Because of its enormous utility and non-invasive nature, faecal hormone monitoring is one of the most powerful tools available in zoo research today.

Age-dependent changes in sperm production, semen quality, and testicular volume in the black-footed ferret (Mustela nigripes).

The black-footed ferret (Mustela nigripes), which was extirpated from its native North American prairie habitat during the 1980s, is being reintroduced to the wild because of a successful captive-breeding program. To enhance propagation, the reproductive biology of this endangered species is being studied intensively. The typical life span of the black-footed ferret is approximately 7 yr. Female fecundity declines after 3 yr of age, but the influence of age on male reproduction is unknown. In this study, testis volume, seminal traits, sperm morphology, and serum testosterone were compared in 116 males from 1 to 7 yr of age living in captivity. Results demonstrated that testes volume during the peak breeding season was similar (P > 0.05) among males 1 to 5 yr of age, reduced (P < 0.05) among males 6 yr of age, and further reduced (P < 0.05) among males 7 yr of age. Motile sperm/ejaculate was similar in males 1 to 6 yr of age but diminished (P < 0.05) in those 7 yr of age. Males at 6 and 7 yr of age produced fewer (P < 0.05) structurally normal sperm than younger counterparts; however, serum testosterone concentrations were not reduced (P > 0.05) in older males. Histological comparison of testicular/epididymal tissue from 5- and 7-yr-old black-footed ferrets confirmed that the interval between these two ages may represent a transitional period to reproductive senescence. In summary, functional reproductive capacity of male black-footed ferrets exceeds that of females by at least 2 yr. Testes and seminal quality are indistinguishable among males 1 to 5 yr of age, with progressive reproductive aging occurring thereafter.

Reproductive inefficiency in male black-footed ferrets (Mustela nigripes).

The black-footed ferret (Mustela nigripes), once considered extinct, has benefited from captive breeding and subsequent reintroduction into native habitat. A high proportion of females (>90%) exhibit estrus in captivity during the spring breeding season. However, many males considered to be prime-breeding age (1-3 years old) fail to sire offspring. Breeding records in 1995 revealed that 40 of 73 males (55%) managed under the Black-Footed Ferret Species Survival Plan did not reproduce, despite being provided opportunity. The present study was conducted to determine the incidence and etiology of male reproductive failure in 1996 and 1997. In 1996, 38 of 69 (55%) 1- to 3-year-old males failed to sire offspring. Likewise, 35 of 60 (58%) males did not reproduce in 1997. Overall, 21% of adult males failed to sire young in three consecutive breeding seasons (1995-1997). Electroejaculate traits (ejaculate volume, sperm concentration, motility, morphology, and acrosomal integrity) from 29 proven breeder males were not different (P > 0.05) from those of 23 males that did not sire young (nonproven breeders). However, six categories of reproductive failure were identified for the 73 prime-breeding age, nonproven males: 1) underdeveloped testes (22%); 2) improper breeding position with the female (25%); 3) excessive aggression toward estrous females (9%); 4) copulation with no sperm detected at postcoital lavage (19%); 5) copulation with sperm in the vaginal lavage but no resulting pregnancy (18%); and 6) copulation with no vaginal lavage performed and no resulting pregnancy (7%). These data indicate that combined behavioral and physiologic factors, but not overall sperm quality, influence reproductive performance in male black-footed ferrets managed in captivity. Zoo Biol 19:517-528, 2000. Copyright 2000 Wiley-Liss, Inc.

Horizontal transmission of feline immunodeficiency virus with semen from seropositive cats.

The AIDS virus of cat species, feline immunodeficiency virus (FIV), has been used extensively as an animal model of HIV-1 infection. This felid lentivirus shares many molecular and biochemical traits with HIV-1 and causes similar immunologic and clinical perturbations, most notably CD4+ cell loss, impaired cell-mediated immunity and increased susceptibility to opportunistic pathogens. Previous reports have shown that FIV is transmitted horizontally by biting and vertically in utero and through nursing. Our objective was to determine whether FIV could be venereally transmitted in domestic cats. In the first experiment, susceptibility of the female reproductive tract to mucosal transmission of the FIV isolate, NCSU1, was demonstrated via intravaginal inoculation with infected cultured cells. We next identified virus in electroejaculates from asymptomatic, chronically FIV-NCSU1-infected, adult males. A fragment of FIV gag provirus DNA was detected by nested polymerase chain reaction (PCR) in nonfractionated seminal cells and in swim-up sperm preparations. Additionally, replication-competent virus was isolated from cell-free seminal plasma and seminal cells by co-cultivation with a feline CD4+ T-cell line. In the third study, queens were artificially inseminated via an intrauterine laparoscopic technique with electroejaculates from FIV-NCSU1-infected males. Of six inseminations carried out with fresh semen, three resulted in infection of queens. Lastly, immunohistochemical studies identified potential virus target cell populations in normal female reproductive tissues. In conclusion, these experiments indicate that FIV infection in domestic cats may provide a unique small animal model of sexual transmission of HIV-1.

Effects of equine chorionic gonadotropin, human chorionic gonadotropin, and laparoscopic artificial insemination on embryo, endocrine, and luteal characteristics in the domestic cat.

The effects of gonadotropin treatment and laparoscopic artificial insemination (AI) on embryo quality, serum progesterone and estradiol concentrations, and luteal progesterone content were examined in the domestic cat. These data were compared to similar historical data reported for naturally estrual, mated queens. All queens in this study (n = 32) were treated with eCG followed by 1) natural breeding (eCG-NB), 2) NB and hCG (eCG-NB-hCG), 3) NB and a sham AI procedure (eCG-NB-sham AI), or 4) hCG and actual AI (eCG-hCG-AI). Queens ovulating in response to treatment were ovariohysterectomized, and oviducts and uteri were flushed to collect embryos. Ovarian structures were recorded, corpora lutea (CL) were excised and evaluated for progesterone content, and serum was analyzed for estradiol-17beta and progesterone. Follicle and CL numbers ranged from 0 to 28 and 2 to 42 per cat, respectively, and treatment means did not differ (p > or = 0.05) among groups. Embryos were recovered from oviducts and uterine horns in all treatment groups, and recovery ranged from 60-96%. Mean embryo number per queen ranged from 8.2 +/- 2.6 to 23.2 +/- 3.8 and did not differ (p > or = 0.05) among groups. However, the proportions of unfertilized oocytes were greater (p < 0.05) for groups treated with hCG and/or artificially inseminated, and the proportion of blastocysts produced (31 of 107, 29.0%) was lower (p < 0.05) in the eCG-hCG-AI group than for any other treatment (range, 59 of 116 [50.9%] to 67 of 116 [57.8%]). Not all queens in each group produced good-quality embryos (eCG-NB, 5 of 5; eCG-NB-hCG, 5 of 8; eCG-NB-sham AI, 2 of 5; and eCG-hCG-AI, 3 of 6). Serum progesterone and estradiol-17beta, and total luteal progesterone per ovary did not differ (p > or = 0.05) among treatments. Compared to historical controls (naturally estrual, mated queens), eCG-NB queens produced > 4 times as many good-quality embryos and blastocysts. Similarly, eCG-hCG-AI-treated queens produced > 4 times the number of oocytes and embryos, although a high proportion of these were poor quality and did not develop to blastocysts. Together, these results indicate that queens treated with eCG are capable of consistently producing many good-quality embryos, at least half of which develop to blastocysts in culture. These data support the use of eCG in felids and suggest that other factors are responsible for reduced pregnancy success and small litter sizes following assisted reproduction.

Sensitivity to exogenous gonadotropins for ovulation induction and laparoscopic artificial insemination in the cheetah and clouded leopard.

Ovarian sensitivity to exogenous gonadotropins was assessed in the cheetah (Acinonyx jubatus) and clouded leopard (Neofelis nebulosa) to help optimize artificial insemination (AI). Eighteen female cheetahs were used on 29 occasions and were given i.m. injections of 100, 200, or 400 IU eCG and 100 or 250 IU hCG 80 h later. Twenty-three female clouded leopards were treated i.m. on 27 occasions with 25, 50, 75, 100, 200, or 400 IU eCG followed 80 h later with 75, 140, or 280 IU hCG. Ovaries were examined laparoscopically at 43-48 h after hCG in cheetahs and 39-50 h in clouded leopards. All gonadotropin dosages stimulated ovarian activity in both species, but ovulation success and corpus luteum (CL) morphology varied (p < 0.05) with treatment. For both species, the highest and intermediate eCG dosages resulted in ovulation in a high proportion (72-100%) of females. The lowest eCG dosage, although capable of stimulating follicular development, compromised ovulation and resulted in few (< 26%) postovulatory females. For each species, small CL (2-4-mm diameter) were observed with the highest and lowest eCG dosage, and large CL (5-8-mm diameter) were associated with intermediate eCG dosages. Aged CL (10-12 mm diameter) were observed in 4 of 23 (17.4%) clouded leopards with no prior male exposure, indicating occasional spontaneous ovulation. Nineteen laparoscopic intrauterine AI procedures were performed in eCG/hCG-treated postovulatory cheetahs. Eighteen AI procedures were conducted in eCG/hCG-treated postovulatory clouded leopards. Six of the 13 cheetahs (46%), all in the 200-IU eCG/100-IU hCG group, became pregnant, in contrast to none of the clouded leopards. This study has revealed differences in ovarian activity in two wild felid species as a result of changes in exogenous gonadotropin dosage. Because of this dose-effect response, this comparative approach is necessary to identify a gonadotropin regimen that can mimic "normalcy." Even then, the relatively high AI success in the cheetah compared to the clouded leopard suggests that factors other than ovarian response can dictate the efficiency of assisted reproduction in this taxon.

Reproductive activity in captive female cheetahs (Acinonyx jubatus) assessed by faecal steroids.

Faecal oestradiol and progestogen metabolite excretion was monitored in adult, female cheetahs (Acinonyx jubatus) (n = 26) for 1-24 months. Increased faecal oestradiol excretion was associated with mating or equine chorionic gonadotrophin (eCG) administration for artificial insemination, whereas increased progestogen metabolites were observed during natural and human chorionic gonadotrophin (hCG)-induced pregnant and nonpregnant luteal phases. On the basis of oestradiol excretory patterns, duration of the oestrous cycle (mean +/- SEM) was 13.6 +/- 1.2 days with high oestradiol concentrations lasting for 4.1 +/- 0.8 days. In non-gonadotrophin-treated cheetahs, 75% showed evidence of oestrous cyclicity; however, none evaluated for 1 year or longer were continuously cyclic. Rather, cyclicity was interrupted by periods of anoestrus, often exceeding several months in duration. These inactive ovarian periods were unrelated to season and were not synchronous among females. Mean duration of gestation (breeding to parturition) was 94.2 +/- 0.5 days, whereas duration of faecal progestogen metabolite excretion during the nonpregnant luteal phase was 51.2 +/- 3.5 days. On the basis of progestogen metabolite evaluations, spontaneous ovulation (non-mating induced) occurred only once in two females (2 of 184 oestrous cycles; 1.1%). Peak eCG-stimulated, preovulatory oestradiol concentrations were similar to those associated with natural oestrus, whereas progestogen metabolite profiles after hCG resembled those during pregnant and nonpregnant luteal phases after natural mating. In summary, results confirm that the cheetah is polyoestrus and ovulation is almost always induced. However, new evidence suggests that many females inexplicably experience periods of anoestrus unrelated to season, while 25% of the cheetahs examined expressed no ovarian activity during the study period.